Pain caused by activation of pain-sensing peripheral neurons ("nociceptors") is a major source of human suffering and economic loss. Activation of nociceptors and the transmission of pain signals to the central nervous system, where they give rise to the conscious perception of pain, requires the coordinated participation of a variety of stimulus- and voltage-gated ion channels. Our long-term goal is the development of pharmaceutical agents for interfering with the functions of ion channels in mammalian nociceptors to ameliorate human pain. We will employ as a source of candidate agents a newly-discovered naturally-occurring combinatorial library of diverse peptide ion channel toxins ("atracotoxins") derived from the venom of Australian funnel-web spiders. To identify members of this library with desirable activity, we will screen them for activity against ion channels known to be important for nociceptor function. The first step in the transduction of painful stimuli-such as extreme heat or cold, noxious chemicals, mechanical injury, or inflammatory mediators-is the activation of stimulus-gated ion channels in the plasma membrane of nociceptors. Once a painful stimulus is transduced by stimulus-gated ion channels into an electrochemical signal, it must be transmitted to the CNS as an electrical signal down the axon of the nociceptor. The transmission of pain signals to the CNS requires activation of voltage-gated ion channels in the endings and axons of nociceptors. We will identify blockers of stimulus- and voltage-gated ion channels in the peripheral terminals of nociceptors by screening atracotoxins against cloned nociceptor ion channels expressed in Xenopus laevis oocytes. Once we have identified a subset of atracotoxins that are active against cloned nociceptor ion channels, we will test each of them against the corresponding native channels in acutely cultured sensory ganglion nociceptors. This secondary screen is essential for excluding from future in vivo studies of candidate analgesics those atracotoxins that are inactive against ion channels in their native state in the nociceptor plasma membrane. These studies will identify lead pharmaceutical agents for use as novel treatments for severe human pain with significant advantages over those that are currently available. In particular, by targeting ion channels present in the peripheral terminals of nociceptors, we open up the possibility for local drug application to the site of peripheral pain. Once these lead agents are identified, we will collaborate with established experts to test these agents in accepted in vivo preclinical mammalian models of pain.